Case Report: Novel compound heterozygous TPRKB variants cause Galloway-Mowat syndrome

Background Galloway-Mowat syndrome (GAMOS) is a rare genetic disease characterized by early-onset nephrotic syndrome and microcephaly with central nervous system abnormalities. Pathogenic variants in genes encoding kinase, endopeptidase, and other proteins of small size (KEOPS) complex subunits cause GAMOS. The subunit TPRKB (TP53RK binding protein) has been reported in only two patients with GAMOS with homozygous missense variants. Clinical report Herein, we described a three-year-old male with GAMOS. He exhibited developmental delay, developmental regression, microcephaly, distinctive facial features, skeletal abnormalities, and epilepsy. Brain magnetic resonance imaging revealed progressive brain atrophy, delayed myelination, T2-hypointense signals in the thalamus, and multiple intracranial abnormal signals on diffusion-weighted imaging. He presented with relapsing nephrotic proteinuria exacerbated by upper respiratory tract infections and progressive renal function decline. Exome sequencing identified compound heterozygous missense and frameshift variants in TPRKB: c.224dup, p.(Ser76IlefsTer3) and c.247C>T, p.(Leu83Phe). Conclusions Our study supports that pathogenic TPRKB variants cause KEOPS complex-related GAMOS.

TPRKB (TP53RK binding protein) (MIM*608680) encodes a subunit of the highly conserved kinase, endopeptidase, and other proteins of small size (KEOPS) complex.The human KEOPS complex comprises OSGEP, TP53RK, TPRKB, LAGE3, and GON7 (9).It regulates the universal chemical modification of tRNA, N 6 -threonylcarbamoyl adenosine (t 6 A), which is essential for normal cell growth and accurate translation by ribosomes (10).The KEOPS complex is involved in human podocyte migration through impaired cell proliferation, increased apoptosis, genomic instability, and defects in actin regulation (4).Biallelic pathogenic variants in genes of the KEOPS complex are responsible for GAMOS in many patients (3,4).Renal biopsies of patients with KEOPS complex-related GAMOS reveals mainly focal segmental glomerulosclerosis or diffuse mesangial sclerosis, with partial podocyte foot process effacement on electron microscopy (4).In contrast, GAMOS 5 (MIM#617731) caused by TPRKB has only been reported in two patients with homozygous missense variants (4).Herein, we report the case of a patient with GAMOS with compound heterozygous missense and frameshift variants in TPRKB.

Clinical report
After 40 weeks of gestation without asphyxia, a Japanese boy was born to nonconsanguineous, healthy parents as their first child.His birth weight was 3,450 g [+1.04 standard deviation (SD)], his length was 51.0 cm (+0.90 SD), and his occipitofrontal circumference was 35.0 cm (+1.25 SD).Distinctive facial features such as widely spaced eyes and pointed chin, pectus excavatum, tapered fingers, esotropia, and muscle hypotonia, were noted (Figures 1A-D).He could hold his head up at five months of age and sit with support at eight months of age.At nine months of age, he babbled but could not maintain a sitting position, even with support.At one year and one month of age, antiepileptic drugs were initiated because of the onset of epileptic spasms and focal to bilateral tonic-clonic seizures.His development regressed, losing interest in toys at one year and three months of age and becoming unable to hold his head up at one year and eight months of age.Tube feeding was initiated because of dysphagia and weight loss at two years and one month of age, followed by gastrostomy at three years of age.
He was diagnosed with proteinuria at six months of age.The spot urine protein/creatinine ratio (UPCR) was 3.33 g/g at one year and one month of age.At 2 years and eight months of age, he was diagnosed with NS due to increased proteinuria (UPCR 17.23 g/g) and hypoalbuminemia (Alb 2.9 g/dl) induced by an acute upper respiratory tract infection.As the infection improved, the proteinuria decreased (UPCR 0.97 g/g).He initiated treatment with valsartan.Subsequently, NS relapsed during respiratory infections, with serum albumin ranging from 2.2 to 3.9 g/dl and UPCR ranging from 0.73 to 17.23 g/g.Serum creatine was low (0.15 mg/dl), while the cystatin C value, which was 0.46 mg/L at 2 years of age, increased to 0.66 mg/L at 3 years of age, suggesting a progressive decline in renal function.Renal biopsy was not performed because of respiratory depression during anesthesia.
Metabolic screenings of blood and cerebrospinal fluid, including amino acids, lactic acid, and pyruvic acid, were unremarkable.Interictal electroencephalogram at one year and eight months of age revealed multifocal spikes with slow background activity.Bilateral visual evoked potentials were absent.Electrocardiography, auditory brainstem response, nerve conduction studies, electroretinography, and echocardiography findings were normal.Radiography revealed no bone abnormalities other than scoliosis and pectus excavatum.Abdominal computed tomography scans of the kidneys, bladder, liver, and spleen revealed no abnormalities.Brain magnetic resonance imaging (MRI) showed a diffuse T2-hypointense signal in the thalamus and numerous characteristic aberrant DWI signals in the white matter, internal capsule, and dorsal part of the pons (Figures 1E-J, L).At one year and eight months of age, there was no T2-hypointensity in the subcortical white matter, suggesting hypomyelination (Figure 1L).The T2-hypointensity in the anterior limb of the internal capsule observed at 10 months of age was indistinct at one year and eight months of age, suggesting demyelination (Figures 1H, L).Progressive atrophy of the cerebral hemispheres and cerebellum was observed (Figures 1K, L).
The last physical examination at three years of age showed a body weight of 12.0 kg (−1.1 SD), height of 97.0 cm (+1.1 SD), and head circumference of 42.5 cm (−4.4 SD).He had a social smile, but his visual tracking was poor.He had constant stridor.Hypotonia of the upper limbs and trunk and spasticity of the lower limbs were noted.His deep tendon reflexes were hyperactive with lower-extremity dominance, and the Babinski reflex and ankle clonus were observed.These findings were suggestive of GAMOS.Progression of neuronal symptoms and repeated worsening of urinary proteinuria due to infectious disease are shown along the timeline in Figure 2.

Ethical compliance
This study was approved by the Institutional Review Board Committee of Hamamatsu University School of Medicine (16-076) and was performed after obtaining written informed consent.Consent to publish identifiable images was obtained from the patient's parents.

Discussion
In this study, we describe the case of a patient with novel compound heterozygous TPRKB variants.Two individuals with  E-G).An axial T2-weighted image shows brain atrophy with frontal white matter dominance and diffuse hypointense signal of the thalamus (H) DWI hyperintensities of the bilateral corona radiata and internal capsule are more distinct (I, J).A sagittal T1-weighted image shows atrophy of cerebellar vermis and thinning of the corpus callosum (K).An axial T2-weighted image reveals progressive brain atrophy and no hypointensity in the subcortical white matter.Hypointensity in the anterior limb of the internal capsule is obscured (L).GAMOS with TPRKB variants have been previously described, and the clinical manifestations of the three cases of TPRKB variants, including our patient, are summarized in Table 1 (4).Microcephaly, global developmental delay, spasticity, and distinctive facial features were common to all three cases.Most patients with GAMOS associated with the KEOPS complex had microcephaly, developmental delay, and distinctive facial features, whereas spasticity was not commonly reported (9/33) (4).This may be because patients with GAMOS with TPRKB variants were relatively older children in whom spasticity was evident due to progression of brain atrophy.In our case, proteinuria was diagnosed earlier than in other patients with TPRKB variants, possibly because of the loss-of-function variant.The age of onset of proteinuria in patients with TPRKB-related GAMOS was later than that in patients with KEOPS complex-related GAMOS other than TPRKB-related GAMOS (median, three months) (4).Furthermore, the median age of death for GAMOS cases with variants of KEOPS complex-encoding genes other than TPRKB was reported to be six months of age.In contrast, one individual with TPRKB variant died at 6.8 years of age, and two cases survived at three and six years.TPRKB-related GAMOS may be milder than GAMOS caused by other KEOPS complexrelated genes.The most frequently observed brain MRI anomalies in the GAMOS associated with the KEOPS complex include cortical and cerebellar atrophy, gyration abnormalities, and myelination defects (4).Abnormal DWI signals, as in our case, have been reported in patients with GAMOS with pathogenic variants of TP53RK and OSGEP, which belong to the KEOPS complex (15, 16).MRI of another case of GAMOS attributed to TP53RK showed diffuse T2-hypointense signals in the thalamus (17).In animal studies, knockout of tprkb in zebrafish larvae recapitulated microcephaly, and Tprkb knockout mouse embryos showed a significant reduction in brain size, cortex length, cortex-midbrain midline length, and cortex width (4).Proteomic profiling of fibroblasts from patients with GAMOS caused by OSGEP variants revealed upregulation of the P2X7 receptor signaling complex pathway (15).In multiple sclerosis, the sustained activation of P2X7 receptors induces oligodendrocyte death, demyelination, neuroinflammatory processes, and neurodegeneration (18).The activation of P2X7 receptors may be involved in the brain phenotype of KEOPS complex-related GAMOS.
Our patient had NS with repeated relapses concurrently with respiratory infections.Increased edema in patients with congenital NS is especially common during infection (19).Relapsing NS with mild persistent proteinuria and relapses of the nephrotic range associated with upper respiratory tract infections have been described in familial cases of the NPHS1 variants encoding nephrin, an essential component of the interpodocytespanning slit diaphragm (20).The molecular mechanism of NS relapse caused by genetic defects remains to be elucidated and requires further investigation.
The patients with KEOPS complex-related GAMOS have severe renal and neurological symptoms and die in early childhood (4).Kidney complications are the direct cause of most GAMOS deaths, and the patients with GAMOS should be screened regularly for proteinuria and renal function and evaluated closely (21).The genetic diagnosis of GAMOS is useful for family planning, and prenatal and preimplantation testing is available after genetic counseling.
In conclusion, we identified a patient with GAMOS and compound heterozygous TPRKB variants who had nephrotic proteinuria, developmental delay, developmental regression, microcephaly, distinctive facial features, skeletal abnormalities, epilepsy, progressive brain atrophy, delayed myelination, T2hypointense signals in the thalamus, and multiple intracranial abnormal signals.This is the first case of a patient with GAMOS associated with a frameshift variant in TPRKB.Further accumulation of cases is necessary to establish the common clinical features in patients with pathogenic TPRKB variants.

FIGURE 1
FIGURE 1 Representative clinical findings.(A-D) Photographs of the patient at ten months of age.Widely spaced eyes, epicanthus, left esotropia, medial sparse eyebrow, pointed chin, and thin upper lip vermilion are observed (A, B).Pectus excavatum (C) and tapered fingers (D) are recognized.(E-L) Brain magnetic resonance imaging (MRI) at ten months of age (E-H) and one year and eight months of age (I-L).Axial diffusion-weighted imaging (DWI) demonstrated elevated signal in the bilateral periventricular white matter, deep and subcortical white matter of the bilateral posterior lobe, posterior limb of the internal capsule, and dorsal part of the pons (E-G).An axial T2-weighted image shows brain atrophy with frontal white matter dominance and diffuse hypointense signal of the thalamus (H) DWI hyperintensities of the bilateral corona radiata and internal capsule are more distinct (I, J).A sagittal T1-weighted image shows atrophy of cerebellar vermis and thinning of the corpus callosum (K).An axial T2-weighted image reveals progressive brain atrophy and no hypointensity in the subcortical white matter.Hypointensity in the anterior limb of the internal capsule is obscured (L).

FIGURE 2
FIGURE 2 Timeline of neurological and renal symptoms.Red allows indicate respiratory infections.Alb, serum albumin; UPCR, spot urine protein/creatinine ratio.

FIGURE 3 TPRKB
FIGURE 3 TPRKB variants in the patients with Galloway-Mowat syndrome.(A) Compound heterozygous variants in TPRKB.Sanger sequencing shows a paternal T duplication (yellow) on the left and a maternal missense variant on the right.(B) Schematic presentation of the TPRKB protein and location of altered residues.Previously reported TPRKB variants are depicted above, and the variants identified in our case are shown below.The predicted secondary structure modified from the PDB (code 6WQX; https://www.rcsb.org/) is indicated for β-strand and α-helix conformation as triangle and zigzag lines, respectively.Multiple amino acid sequences of TPRKB were aligned using the ClustalW tool (http://www.genome.jp/tools/clustalw).